JP5343801B2 - Heat transfer device for gene processing device and gene processing device - Google Patents

Description

  TECHNICAL FIELD The present invention relates to a heat transfer device for transferring heat to an object and a device provided with a heat transfer mechanism, and in particular, a heat transfer device for a reaction vessel used for genetic testing or the like and a gene processing device provided with a heat transfer mechanism. About.

  Genetic testing detects the difference in genes that cause individual differences in drug metabolism, such as single nucleotide polymorphisms (SNPs), thereby avoiding side effects and providing optimal medical care for patients ( In recent years, this technology has attracted attention as a technology that can be used for tailor-made medical services that can provide drugs.

  In the genetic testing technology, models that perform testing using nucleic acid (DNA) purified from a specimen have been introduced into various markets from testing equipment companies. These devices require a temperature control device for gene amplification reactions. Generally, a reaction vessel such as a plate or tube is set in a heat transfer block that matches the shape of each tip, and pressed from above with a lid. Heat transfer is carried out, and gene amplification reaction is carried out.

  However, it is difficult to completely match the tip shape of these reaction vessels with the shape of the heat transfer block, and there is a problem that the gene amplification reaction cannot be performed efficiently. In particular, when the reaction part is made of metal instead of resin as in Patent Document 1, the adhesion to the heat transfer block due to expansion due to an increase in internal pressure during gene amplification reaction like a plate or tube made of resin. However, it was not possible to expect to improve the temperature, and how to improve the adhesion between the reaction vessel formed of metal and the heat transfer block was a big problem.

  On the other hand, in a heat transfer device used for heat dissipation and heat conduction of an electronic module, a low melting point metal is arranged at a contact portion with an object of the heat transfer device, and heat transfer is achieved by melting the low melting point metal by heat. There is something improved. Patent document 2 is applied between a microprocessor and a chip and a heat sink. Patent Document 3 is applied between a thermoelectric power generation module and a heat sink. These combinations are used in a state where contact is maintained once set. In both Patent Documents 2 and 3, the contact surface of the object and the heat transfer device is a flat contact.

  In the conventional heat transfer device using the low melting point metal as described above, there has been no use mode in which contact and release with the object are repeatedly performed. Furthermore, there has been a demand for a heat transfer device that is used for genetic testing or the like and has good adhesion to a container having a non-planar temperature control portion.

International Publication Number WO2009 / 054473 JP 2007-173825 A JP 2007-1116086 A

  From the background as described above, an object of the present invention is to provide a heat transfer device in which a temperature control unit used for genetic testing or the like has good adhesion and thermal conductivity to a non-planar container. In addition, without depending on the variation in the processing dimensions of the gene analysis chip, the gene analysis chip having an uneven surface is closely contacted with the heat transfer portion that performs the gene amplification reaction without any gaps, and the gene amplification reaction is performed. Thus, a gene processing apparatus with improved gene amplification reaction reactivity is provided.

In response to this problem, the present invention arranges a low melting point metal having a melting point of 99 ° C. or less at a position where the reaction well formed of metal contacts the heat transfer block, so that the low melting point metal melts during the gene amplification reaction. This is solved by improving the adhesion between the reaction well formed of metal and the low melting point metal. In particular,
As a first invention, a heat transfer device for a gene processing apparatus that conducts heat with a protruding portion of an object , wherein the object is an analysis chip composed of a substrate and a cover material, and corresponds to the protruding portion And a low melting point metal layer formed in the recess, and the melting point of the low melting point metal of the low melting point metal layer is greater than 25 ° C and 99 ° C or less. This is a heat transfer device for a certain gene processing device .
Further, as a second invention, in the first invention, the heat transfer block is a heat transfer device for a gene processing device, which is any one of silver, copper, and aluminum.
Furthermore, as a third invention, in the first or second invention, the low melting point metal of the low melting point metal layer has a melting point of the low melting point metal that is outside the temperature cycle range of the gene amplification reaction. .
Furthermore, as a fourth invention, in the first to third inventions, the material for the low melting point metal layer is a heat transfer device for a gene processing device, which is an alloy containing lead, tin, cadmium, bismuth, or indium. is there.
Further, as a fifth invention, in the first to fourth inventions, the target object is plate-shaped and has a plurality of protruding portions on the plate surface, and a plurality of the recesses corresponding to the gene processing are formed. This is a heat transfer device for a device.
Furthermore, as a sixth invention, in the first to fifth inventions, a heat transfer device for a gene processing apparatus having a thermo module thermally connected to a heat transfer block is provided.
Furthermore, as a seventh invention, a gene processing apparatus comprising the heat transfer device for a gene processing apparatus according to the first to sixth inventions is provided.

  According to the present invention, if the melting point of the low melting point metal is higher than the melting point of the low melting point metal during the heat treatment, the low melting point metal is melted, so that it is possible to contact the gap between the object and the heat transfer part, thereby improving the thermal conductivity. it can. In addition, since the low melting point metal layer exists between the concave part of the heat transfer block corresponding to the projecting part (convex part) of the target object and the convex part, it is possible to make contact without depending on the dimension of the convex part. become.

  By using such a heat transfer device for a gene processing device, the entire surface of the gene analysis chip that contacts the heat transfer portion can be brought into close contact, so that the speed of the gene amplification reaction can be increased.

It is a cross-sectional schematic diagram of the Example for demonstrating the heat-transfer apparatus of this invention. It is a cross-sectional schematic diagram of the comparative example for demonstrating the heat-transfer apparatus of this invention. It is a perspective view of a gene analysis chip. It is a comparison figure showing the temperature change of the Example and comparative example in the gene amplification reaction by PCR method. It is a comparison figure of the FAM fluorescence intensity in the invader reaction using the gene processing apparatus of an Example and a comparative example. It is a comparison figure of the RED fluorescence intensity in the invader reaction using the gene processing apparatus of an Example and a comparative example.

  FIG. 1 is a schematic cross-sectional view of an embodiment of a gene processing apparatus using the heat transfer device of the present invention. The heat transfer device of the present invention is a heat transfer mechanism for performing heat treatment such as heating and cooling on a predetermined position of an object provided with a heat transfer block in contact with the object. In the example of the heat transfer device 100 in FIG. 1, the reaction vessel 2 formed on the gene analysis chip 1 is a target of heat treatment, and the protruding bottom portion of the reaction vessel (substrate protruding portion 8) is a portion that conducts heat. The heat transfer device of the present invention has a recess 6 corresponding to the protruding portion on the surface in contact with the object of the heat transfer block, and a low melting point metal layer 14 is formed in the recess. According to the heat transfer device of the present invention, by setting the temperature higher than the melting point of the low melting point metal used for the low melting point metal layer during the heat treatment, the projecting portion of the object disposed in the recess and the gap between the heat transfer blocks are reduced. It becomes possible to fill with liquid metal. For this reason, thermal conductivity can be improved and the precision of temperature control can be raised.

  FIG. 2 is a schematic cross-sectional view of an embodiment of a gene processing apparatus using a conventional heat transfer apparatus shown as a comparative example. Some reference numerals are omitted for the same parts as in FIG. In the heat transfer device 200 of the comparative example, heat transfer is performed by bringing the heat transfer block 9 into direct contact with a portion that conducts heat. However, in this method, since it is necessary to ensure the variation in the processing dimension of the convex portion 8 on the contact side with the heat transfer device of the target object and the alignment margin between the target convex portion and the heat transfer block concave portion 6, It is difficult to bring the convex portion (here hemispherical) into contact with the concave portion of the heat transfer block corresponding to the convex portion, and there is a problem that a slight gap is generated and the efficiency of heat conduction is lowered.

  As shown in FIG. 1, in the heat transfer device of the present invention, since the recess 6 of the heat transfer block can be made one size larger than the shape of the corresponding object protrusion 8, the alignment margin can be increased. it can. Moreover, since it is not necessary to make the shape of a recessed part exactly correspond to the shape of a convex part, the cross-section 6a with a circular arc shape, the rectangular recessed part 6b, etc. can be selected suitably, for example. As a material for the heat transfer block, a material having a high thermal conductivity generally used as a heat spreader can be used, and specifically, a metal such as aluminum, silver, or copper can be preferably used.

  As a material of the low melting point metal layer 14 formed in the recess 6 of the heat transfer block, an alloy containing lead, tin, cadmium, bismuth, or indium can be suitably used. The low melting point metal may be solid at room temperature (25 ° C.) and liquid at the heat treatment temperature.

  The shape of the low melting point metal layer may be formed along the recess or may be deposited on the bottom of the recess. When at least the low melting point metal layer is melted and liquefied, and the object is placed on the heat transfer block, an amount of the low melting point metal that is in contact with the protruding recess of the object and spreads in the recess is deposited.

  In the heat transfer device of the present invention, the thermo module 16 is disposed on the opposite surface of the contact surface between the heat transfer section block 9 and the object, and is thermally connected to the heat transfer block, thereby the object (reaction vessel 2). The temperature can be controlled. Furthermore, the radiator 17 can be efficiently cooled by being disposed in a region (for example, the opposite surface) other than the portion where the heat transfer block of the thermo module is disposed and thermally connected to the thermo module. Note that the term “thermally connected” means a state in which a temperature change can be given to at least one of the two by heat exchange.

  Next, the case where the heat transfer apparatus of this invention is used for a gene processing apparatus is demonstrated. As described in the background art, the gene processing apparatus in the present invention is an apparatus used when performing heat treatment in genetic testing or the like, and is used for realizing a thermal cycle for nucleic acid amplification by gene amplification reaction, for example. Can do. Examples of gene amplification reactions include PCR, NASBA, SMAP, and LCR methods. In the case of the PCR method, a cycle in which a denaturation temperature of about 95 ° C. and an annealing temperature of 60 to 70 ° C. are alternately repeated is performed in the gene processing apparatus.

  When used in a gene processing apparatus, the heating temperature for melting the low melting point metal layer 14 needs to be a temperature that does not inhibit the reaction. For example, the temperature is such that the enzyme is not inactivated, and in the case of the PCR method, it is preferably 99 ° C. or lower. That is, the low melting point metal used preferably has a melting point of 25 ° C. or higher and 99 ° C. or lower. Furthermore, when performing temperature control having a temperature cycle in a certain temperature range as in the PCR method, the melting point of the low melting point metal is more preferably outside the temperature cycle range. This is because, when the melting point of the low melting point metal is in the temperature cycle range, the low melting point metal is affected by the latent heat accompanying the phase change and the solid and liquid of the low melting point metal, thereby hindering the speeding up of the temperature cycle. For example, in the case of a temperature cycle (see FIG. 4) of a gene amplification reaction by the PCR method in Examples described later, temperature control is performed in the range of 68 ° C. to 95 ° C. In this case, the melting point is preferably 68 ° C. or lower, or 95 ° C. or higher.

  A perspective view of the gene analysis chip 1 used in the gene processing apparatus of the present invention is shown in FIG. 1 is a cross-sectional view taken along line A-A ′ of FIG. 3. In the gene analysis chip of FIG. 3, a plurality of reaction containers 2 are provided on a plate-like substrate, and the reaction reagent 13 in the reaction container and the fixed reagent 11 are mixed and reacted by heat melting of the sealing material 12. Detection can be performed in each reaction vessel. In the gene analysis chip of FIG. 3, each reaction container is formed by bonding the base material and the cover material so that the convex portion 8 of the base material 7 and the concave portion of the cover material 5 are matched. At this time, a sealing material that covers the fixed reagent and the fixed reagent is formed in advance in each concave portion of the cover material. In addition, the base material is provided with a flow path 15 that communicates with each convex portion, and a reaction reagent can be injected from the sample injection port 3 to distribute the reaction reagent to each reaction container.

  According to the gene processing apparatus of the present invention, the entire surface (the back surface of the base material 7) in contact with the heat transfer portion of the gene analysis chip can be brought into close contact, so that the speed of the gene amplification reaction can be increased. Further, since the temperature rises to a temperature equal to or higher than the low melting point metal during the gene amplification reaction, the analysis chip and the heat transfer portion can be in close contact without depending on the size of the convex portion of the analysis chip.

  Hereinafter, experimental results of gene testing will be shown using the gene processing apparatus (Example) of the present invention shown in FIG. 1 and the gene processing apparatus of the comparative example shown in FIG.

  First, the fixing reagent 11 was placed in the recess 6 of the cover material 5. As the fixing reagent 11, allele probe 1, allele probe 2, invader probe and FRET probe 1, FRET probe 2, and Cleavease (registered trademark) used for Invader reaction (registered trademark) were placed and dried by heating. Allele probe 1 and allele probe 2 are labeled with FAM fluorescence and RED fluorescence, respectively.

  Next, EMW-0003 manufactured by Nippon Seiki Co., Ltd. was introduced as the sealant 12 into all the recesses 6 of the cover material 5. In addition, the injection amount of the sealing agent 12 with respect to the recessed part 6 of one place is 3.5 microliters. The sealing agent 12 was heated to 80 to 100 ° C., dispensed into the recess 6 in a melted state, centrifuged with a plate centrifuge, and then solidified again at room temperature. Thereby, the reaction container 2 in FIG. 1 covers the fixing reagent 11 with the sealant 12.

  Next, using the heat seal tester manufactured by Tester Sangyo Co., Ltd., the cover material 5 and the substrate 7 produced as described above are heated under the conditions of one-side heating from the substrate 7 side at 220 ° C., 0.5 MPa, 2.2 seconds. And bonded by heat sealing.

  Next, a mixed solution of purified genomic DNA, invader buffer, and water for diluting it was supplied as a reaction reagent. The reaction reagent 13 was distributed from the sample inlet 3 of the cover material 5 to all the reaction containers 2 through the flow path 15. Note that the supply amount of the reaction reagent 13 to one reaction vessel 2 is 12 to 15 μL. The gene analysis chip 1 was produced by the above process.

  Thereafter, the gene analysis chip 1 was set in the gene processing apparatus according to the example of the present invention and the gene processing apparatus as a comparative example. First, in the apparatus, a part of the flow path 15 was crushed by an external force so as to prevent the reaction liquid during the reaction from passing between the reaction containers 2, thereby sealing the reaction containers 2. At that time, heat was applied simultaneously with the external force, and the cover material 5 and the sealant layer of the substrate 7 were thermally welded to perform stronger sealing. Sealing was performed under the conditions of 170 ° C., 240 kgf, and 1.8 seconds.

  Next, the gene analysis chip 1 was heated from the substrate 7 side and the cover material 5 side in the apparatus, and nucleic acid amplification was performed by the PCR method. In this example, a low melting point metal having a melting point of 95 ° C. was used. First, the sealing agent 12 was melted by heating at 95 ° C. for 2 minutes, which is an initial denaturing condition, and the fixing reagent 11 and the reaction reagent 13 were brought into contact with each other. At this time, the low melting point metal that is in contact with the substrate 7 side is also melted, thereby filling the gap between the substrate protrusion 8 and the aluminum block 9. In this state, a gene amplification reaction was performed. As setting conditions for the gene amplification reaction, 30 cycles of 95 ° C., 25 seconds holding and 68 ° C., 35 seconds holding were performed for 1 cycle, and then Cleaase was inactivated at 99 ° C. for 2 minutes. Thereafter, the temperature was lowered to 60 ° C., and while performing the invader reaction, the fluorescent substance in the reaction vessel 2 was detected once every 30 seconds, and the state of the reaction for each hour was observed.

  FIG. 4 is a comparison of the set temperature of the thermo module 16 in the PCR reaction step and the temperature change in the reaction vessel in the examples and comparative examples. As indicated by the arrows, when the apparatus of the example is used, the set temperature change is closer than in the comparative example. That is, it can be seen that the efficiency of heat conduction is improved.

  5 and 6 show the results of fluorescence detection. FIG. 5 shows the time change of the FAM fluorescence intensity when the reaction is carried out using the apparatuses of the example and the comparative example. Compared with the comparative example, it can be seen that the reactivity (rising of the fluorescence intensity) was improved by performing the reaction in the gene processing apparatus equipped with the heat transfer device of the present invention. FIG. 6 shows temporal changes in the RED fluorescence intensity when the reaction is performed using the apparatuses of the example and the comparative example. It can be seen that also in the RED fluorescence intensity, the reactivity was improved as compared with the comparative example.

DESCRIPTION OF SYMBOLS 1 ... Gene analysis chip 2 ... Reaction container 3 ... Sample injection port 4 ... Cover 5 ... Cover material 6 ... Concave part 7 ... Substrate 8 ... Substrate convex part 9 ... Heat-transfer block 10 ... Gap 11 ... Fixing reagent 12 ... Sealing agent 13 ... reaction reagent 14 ... low melting point metal 15 ... flow path 16 ... thermo module 17 ... radiator (heat sink)
100 ... Heat Transfer Device 200 ... Heat Transfer Device (Comparative Example)

Claims (7)

A heat transfer device for a gene processing device that conducts heat with a protruding part of an object,
The object is an analysis chip composed of a substrate and a cover material,
A heat transfer block having a recess provided corresponding to the protruding portion;
A low melting point metal layer formed in the recess,
Equipped with a,
The melting point of the low melting point metal of the low melting point metal layer is greater than 25 ° C. and 99 ° C. or less.
A heat transfer device for a gene processing device. The heat transfer device for a gene processing device according to claim 1, wherein the heat transfer block is one of silver, copper, and aluminum. The heat transfer device for a gene processing device according to claim 1 or 2 , wherein the melting point of the low melting point metal in the low melting point metal layer is outside the temperature cycle range of the gene amplification reaction. The heat transfer device for a gene processing device according to any one of claims 1 to 3 , wherein the material of the low melting point metal layer is an alloy containing lead, tin, cadmium, bismuth, or indium. The object is a plate-shaped, has a plurality of protruding portions on the plate surface, according to any one of claims 1 to 4, wherein a plurality of said recesses corresponding thereto are formed Heat transfer device for gene processing equipment . The heat transfer device for a gene processing device according to any one of claims 1 to 5 , further comprising a thermo module thermally connected to the heat transfer block. A gene processing apparatus comprising the heat transfer device for a gene processing apparatus according to any one of claims 1 to 6 .